KR101465866B1 - Nanocomposite of Biocompatible PHEMA derivatives/Ag having antibacterial and antifouling activity - Google Patents

Abstract

The present invention discloses a polymer-silver nanocomposite comprising a poly (2- (2-acetoxyacetoxy) ethyl methacrylate and silver as a polymer and a preparation method thereof. The biocompatible polymer-silver nanocomposite of the present invention has antibacterial, antifouling And biocompatibility, and can be used for various purposes such as biomedical materials, coating materials, and films that require antibacterial and antifouling, and the manufacturing process also does not require a separate crosslinking process, thereby increasing the convenience of the process.

Description

 The present invention relates to a biocompatible PHEMA derivative / antifouling activity / biocompatible PHEMA derivative having antibacterial and antifouling properties,

This invention relates to biocompatible PHEMA derivatives / silver nanocomposites and methods for their preparation.

In industry and medicine, the contamination or infection of the bacteria with the material may not only lower the efficacy of the product but also pose a life threat. Bacteria tend to form clusters attached to the surface of industrial materials or biomaterials to form biomembranes easily. These bacterial biofilms are frequently found in sink sinks and heat transfer equipment, and are also found in biomedical materials such as food processing systems or implants. For this reason, prevention of bacterial surface adhesion with bacterial elimination is recognized as a very important task in industrial and medical aspects.

Therefore, many studies have been carried out to prevent surface adhesion of bacteria. The first is a method of fixing an antifouling material such as polyethylene glycol (PEG) to the surface. However, this is effective in blocking bacterial surface adhesion, but there is a limit to the ongoing aspect because it is not possible to remove the biological film already made by the bacteria attached to the surface. The second is to introduce antibiotics or metals such as quaternary ammonium compounds (QAC) onto the surface. But even with this method, antibiotics can kill bacteria, but they can not prevent bacterial adhesion. That is, the above methods do not simultaneously have antifouling property and antimicrobial property.

There has been an attempt to secure antifouling property and antimicrobial property by using a biocompatible polymer used for industrial materials or biomaterials.

Tissue cell polystyrene (TCPS) is widely used as a biocompatible material, but it has excellent cell adhesion and proliferation, but it is deficient in preventing bacterial adhesion.

PHEMA (poly (2-hydroxyethyl methacrylate)) is a polymer having excellent biocompatibility, antibacterial and cytotoxic properties, but it has disadvantages that adhesion and proliferation of cells are limited. In addition, since PHEMA has a hydroxyl group at the side chain of the polymer side chain, it has a high hydrophilicity and thus has poor film forming properties and has a very poor long-term stability in water. In order to increase the stability, although the stability is increased by further performing the crosslinking process, there is a disadvantage that a separate crosslinking process is further required.

On the other hand, silver nanoparticles can be used to deactivate a wide variety of microorganisms as antimicrobial materials widely used in the industry. However, since silver nanoparticles have a problem of being toxic to cells, it is required to eliminate or reduce them.

Korean Patent Publication No. 2011-0017180 discloses a curable resin composition containing nano-sized silver particles to impart antimicrobial and bactericidal properties and to improve durability.

Korean Patent Laid-Open Publication No. 2009-0074010 discloses a coating composition for a steel pipe having a function of antibacterial function including silver nano and a method for producing the same.

Therefore, in order to realize a film which has low cytotoxicity and antibacterial and antifouling properties and can prevent the adhesion of bacteria, it is necessary to invent a composite material in which silver nanoparticles having a proper concentration are applied to a biocompatible polymer. Particularly biocompatible polymer / silver nanocomposites having antimicrobial properties, antifouling properties and biocompatibility at the same time are not disclosed.

The present invention is directed to a biocompatible polymer / silver nanocomposite having antimicrobial, antifouling and biocompatibility and a method for producing the same.

In one embodiment, the present invention provides a polymer-silver nanocomposite comprising a poly (2- (2-acetoxyacetoxy) ethyl methacrylate and silver as a polymer. In one embodiment, the polymer comprises about 85 to 99 wt% The silver, which may be used herein, in one embodiment, is reduced silver, which is incorporated into the polymer.

In another aspect, the present disclosure provides a method of forming a coating comprising coating a solution of a biocompatible polymer, poly (2- (2-acetoxyacetoxy) ethyl methacrylate and a silver compound, on a substrate and irradiating the coated mixture with an electron beam, Silver nanocomposite according to claim 1, comprising the step of reducing the silver compound.

In one embodiment according to the present application, the silver compound is perchloric acid, silver nitrate, or silver carbonate.

In one embodiment according to the present application, light may be near ultraviolet, ultraviolet or gamma rays.

In one embodiment according to the method of the present invention, the heat treatment is performed at a temperature of about 80 캜 to 120 캜.

In another aspect, the present invention also provides an antimicrobial, antifouling, and biocompatible polymer film produced using the biocompatible polymer-silver nanocomposite according to the present invention.

In another aspect, the disclosure also provides a coating material having antimicrobial, antifouling, and biocompatibility, prepared using the biocompatible polymer-silver nanocomposite according to the present invention.

In another aspect, the present invention also provides a biomedical material having antimicrobial, antifouling and biocompatibility prepared using the biocompatible polymer-silver nanocomposite according to the present invention.

The biocompatible polymer / silver nanocomposite of the present application is excellent in antibacterial, antifouling and biocompatibility, and has increased stability even in water, and can be widely used as a biomedical material and the like.

In addition, the biocompatible polymer / silver nanocomposite of the present invention does not require a separate crosslinking step, thereby increasing convenience in terms of the manufacturing process. The PHEMAAA used in the present invention serves to prevent aggregation of the reduced silver nanoparticles and thus has surface stability without any additional crosslinking process.

In addition, the dispersibility of the silver nanoparticles in the composite is also excellent, so that the antibacterial properties are exhibited properly.

1 is a schematic diagram of the PHEMAAA / silver nanocomposites of Examples 1 and 2 according to one embodiment of the present invention.
FIG. 2 is a photograph showing the PHEMAAA / perchloric acid mixture (a) before the reduction process, the PHEMAAA / silver nano composite (b) subjected to ultraviolet (UV) reduction according to one embodiment of the present invention, c) was confirmed by FE-SEM (field emission scanning electron microscopy).
FIG. 3 is a graph showing the relationship between the PHEMAAA / perchloric acid mixture before the reduction process, the PHEMAAA / silver nanocomposite subjected to ultraviolet (UV) reduction according to one embodiment of the present invention and the PHEMAAA / XPS spectral results obtained by measuring the surface state of the film.
Figures 4a and 4b are graphs showing the results for P. aeruginosa (4a) and S. epidermidis (ATCC 12228) (4b) attached to PS (polystyrene), PHEMAAA and PHEMAAA / As a result of photographing the effect with CLSM, green and red represent living bacteria and killed bacteria, respectively.
Figure 5 shows the results of measuring the antimicrobial effect of PHEMAAA and the PHEMAAA / silver nanocomposite (UV treatment) according to one embodiment of the present invention on P. aeruginosa and Staphylococcus aureus.
FIG. 6 shows the mitochondrial metabolic activity (a) obtained by the MTT assay of TCPS, PHEMAAA and PHEMAAA / silver nanocomposite (UV treatment) according to one embodiment of the present invention, and the cytotoxic activity of human dermal cells obtained from the neutral analysis ).
Figure 7 shows the results of TUNEL analysis of TCPS, PHEMAAA and PHEMAAA / silver nanocomposite (UV treatment) according to one embodiment of the present invention.
Figure 8 shows quantitative analysis results of cell growth of TCPS, PHEMAAA and PHEMAAA / silver nanocomposite (UV treatment) according to one embodiment of the present invention. TCPS (tissue culture polystyrene).

The present disclosure is based on the discovery that the biocompatible polymer / silver nanocomposites herein have both antimicrobial, antifouling, biocompatibility and stability.

In one embodiment, the present invention relates to a biocompatible polymer / silver nanocomposite comprising 85 to 99% by weight of a biocompatible polymer and 0.001 to 5% by weight of silver nanoparticles.

The term " biocompatibility " as used herein means the property of a material capable of coexisting with a living body while performing its original functions without exhibiting adverse effects on the living body over a long period of time.

The biocompatible polymers included herein are synthetic polymers selected from the group consisting of polyacrylic acid, polyacrylic acid derivatives, or combinations thereof. In one embodiment according to the present application, biocompatible polymers include polyacrylic acid derivatives such as ester derivatives, amide derivatives and the like. In one embodiment according to the present application, poly (2- (2-acetoxyacetoxy) ethyl methacrylate (PHEMAAA) is used.

Poly (2- (2-acetoxyacetoxy) ethyl methacrylate) according to the present invention is a derivative of poly (2-hydroxyethyl methacrylate) (PHEMA) having an ester group, ≪ / RTI > acrylate) and acetoxyacetyl chloride. Has lower hydrophilicity and lower glass transition temperature than poly (2-hydroxyethyl methacrylate). In the case of PHEMA, PHEMAAA according to the present invention has a low hydrophilicity and a low glass transition temperature, and has a low hydrophilicity and poor stability in water due to the presence of -OH group at the side chain of the polymer side chain. This is excellent.

The 'silver' particles of the present invention have antimicrobial properties and are free from silver compounds and become a complex with the biocompatible polymer. Silver ions are reduced and used by ultraviolet rays or heat treatment.

As used herein, " composite " means that two or more materials are associated by various chemical and / or physical bonds. In one embodiment according to the present disclosure, the silver of the metal is embedded in a polymer matrix, i.e., poly (2- (2-acetoxyacetoxy) ethyl methacrylate, as shown in FIG.

As used herein, the term " nanocomposite " refers to a complex having at least one of two or more kinds of materials composing the complex having nanoscale dimensions, and the definition can be referenced as follows: The International Union of Pure and Applied Chemistry & . In one embodiment according to the present invention, the size of the nanoparticles is not limited to about 1 to about 100 nm, especially about 7 to about 100 nm, more particularly about 1 to about 50 nm, even more particularly about 1 to 25 nm .

In the biocompatible polymer / silver nanocomposite of the present application, the biocompatible polymer is about 85 to 99% by weight, preferably about 90 to 96% by weight based on the total complex. The content of the silver particles is 0.001 to 5% by weight.

In another aspect, the present invention relates to a method of directly "in situ" manufacturing metal nanoparticles in a polymer. (Step 1) of coating the biocompatible polymer and silver mixture solution on a substrate including a substrate, for example, glass, silicon, various polymer surfaces, fibers, paper, and metal, Silver nanocomposite, comprising the step of irradiating or heat-treating silver (step 2).

The coating in step 1 above may be carried out using various known methods. For example, horizontal spin casting may be used. In the horizontal centrifugal casting method, a mixture is put into a rotating cylindrical mold and is rotated at an appropriate speed, for example, about 300 to 3000 rpm to mold it by centrifugal force. The rotation time can be appropriately adjusted and can be rotated for about 10 seconds to 5 minutes, preferably about 20 seconds to 1 minute. The 'silver compounds' that can be used include salts and colloidal phases, and silver acetate, perchloric acid silver and the like can be used. As the solvent used in the mixture solution, an inert solvent may be used. For example, THF (tetrahydrofuran) may be used.

The reduction reaction in the step 2 can be generally carried out in two ways. One may use various optical energy, for example, near-infrared, near-ultraviolet, ultraviolet, visible, gamma, or electron beams, preferably ultraviolet. The light energy can be irradiated for a suitable time, for example from about 2 to 10 hours, and the irradiation can take place at about 5 to 20 cm away. The other is a reduction reaction using heat treatment. The heat treatment temperature may be from 90 占 폚 to 99 占 폚.

By this reduction reaction, monovalent silver (Ag ⁺ ) is reduced (Ag ⁰ ).

In another aspect, the present invention is directed to a polymeric film having antimicrobial, bactericidal and biocompatibility properties, prepared using the biocompatible polymer / silver nanocomposite. The polymer film refers to a non-fibrous plate-shaped plastic molding having a thickness of 0.25 mm (1/100 inch) or less.

In another embodiment, the present invention is directed to a coating material having antimicrobial, bactericidal and biocompatibility properties, prepared using the biocompatible polymer / silver nanocomposite.

In another aspect, the present invention relates to a biomedical material having antimicrobial, bactericide and biocompatibility, prepared by mixing the biocompatible polymer / silver nanocomposite. The term 'biomedical material' refers to a substance used for medical purposes designed to perform a desired function without a biological reaction or biological reaction when it comes into contact with blood, body fluids or living tissues in and / or out of a living body. Narrowly refers to a material that can be implanted in a living body to replace or complement the function of the living body without the interaction of the living body. Implant materials, biosensors, tissue engineering, etc.

Hereinafter, embodiments are provided to facilitate understanding of the present invention. However, the following examples are provided only for the purpose of easier understanding of the present invention, and the present invention is not limited to the following examples.

Example 1 shows the preparation of a nanocomposite

PHEMAAA was prepared as follows. PHEMA (Poly (2-hydroxyethyl methacrylate)) was dissolved in pyridine. 1.5 equivalents of acetoxyacetyl chloride to PHEMA was dissolved in THF (tetrahydrofuran), and the mixture was added dropwise thereto, followed by reaction at 50 DEG C for 12 hours. After washing in 1 ml of 50% methanol, it was dissolved in chloroform and precipitated in methanol.

First, the silicon wafer was washed with a piranha solution (70 vol.% Sulfuric acid and 30 vol.% Hydrogen peroxide) and dried in a nitrogen stream for 3 hours. Then, 95% of PHEMAAA and 5% of perchloric acid were mixed to prepare a mixture, which was dissolved in THF at 1% by weight. This was sprayed on a silicon wafer at 3000 rpm for 30 seconds using a horizontal spin casting method. The film thus produced was dried in a vacuum state at room temperature for 12 hours.

Then, the produced film was reduced by the following two methods of ultraviolet ray irradiation and heat treatment. In the first method, the PHEMAAA / perchloric acid mixture was reduced by irradiating ultraviolet rays 9 cm away from the produced film for 3 hours to obtain a PHEMAAA / silver nanocomposite.

In the second method, the PHEMAAA / perchloric acid mixture was reduced by standing the film at 95 ℃ for 3 hours to obtain a PHEMAAA / silver nanocomposite.

Next, the morphology of the PHEMAAA / perchloric acid mixture and the PHEMAAA / nano composite prepared as described above was confirmed by FE-SEM (field emission scanning electron microscopy, SUPRA 55VP (Carl Zeiss, Germany)). As a result, as shown in FIG. 2 (a), PHEMAAA / perchloric acid showed a homogeneous distribution of nanoparticles while self-reduction or clustering of perchloric acid occurred despite the reduction treatment. On the other hand, as shown in FIG. 2 (b), the PHEMAAA / silver nanofilm showed uniform silver nanoparticles uniformly dispersed on the surface without aggregation. Also, as shown in Fig. 2 (c), it was found that the surface of the film remained stable without being damaged even in water, and the stability in water was also confirmed.

For more detailed surface analysis, the surface state of the film was measured with XPS (X-ray photoelectron spectroscopy, Sigma Probe, Thermo Scientific, UK). The graph of FIG. 3 shows the comparison of PHEMAAA / silver nanocomposite (a) before reduction, PHEMAAA / silver nanocomposite (b) reduced by UV for 3 hours and PHEMAAA / silver nanocomposite (c) reduced by heat treatment for 3 hours It is an XPS spectrum. The 3d peak shift of the silver nanoparticles and the binding energy of the monovalent silver (Ag ⁺ ) and silver (Ag ⁰ ) surfaces were compared. First, we can see that the mixture of PHEMAAA / perchloric acid has a higher binding energy of 358.5 eV, 358.4 eV at 368.2 eV when comparing the binding energy at the silver 3d5 / 2 peak of two PHEMAAA / silver nanocomposite films subjected to ultraviolet or thermal reduction treatment . This proves that Ag ⁺ is reduced to Ag ⁰ . Would also appear to 3d _5/2 peak and 3d3 / 2 peak gap between 6eV also shows a Ag ^0. The remaining chloride ions may have played a role in blocking the bacteria, but no peak for chloride ions.

Experimental Example 1. Bacterial adhesion or antifouling investigation

In order to confirm that the PHEMAAA / silver nanocomposite of the present invention has the property of preventing the bacteria from adhering to the surface, the live / dead bacterial viability test was applied to Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcus aureus (1 × 10 ⁸ , 5 × 10 ⁷ , and 1 × 10 ⁸ CFU), respectively. Specifically, the above three bacteria were cultured in LB (Luria? Bertani) for 12 hours at 37 占 폚, centrifuged and washed with PBS. Each of the bacteria was then diluted to 1 × 10 ⁸ , 5 × 10 ⁷ , and 1 × 10 ⁸ CFU (colony forming unit) / mL. Three samples as described in Fig. 4 were coated on 1.2 cm x 1.2 cm silicon wafers and immersed in 2.5 ml of each bacterial suspension for 24 hours at 50 rpm and at 25 占 폚. After a day, the specimens were carefully washed with third distilled water, and 1.5 μL of BacLight Live / Dead bacterial viability kit (L7012, Molecular probes, USA) was diluted with 1 ml of distilled water and each bacteria was stained for 15 minutes under dark conditions . Live bacteria are green, dead bacteria are red. The surface images were confirmed by confocal laser scanning microscopy (CLSM, Eclipse 90i, Nikon, Japan).

The results are shown in Figures 4a and b. Figure 4 shows CLSM values for Pseudomonas aeruginosa, Staphylococcus epidermidis and Staphylococcus aureus attached to PS (poly styrene negative control), PHEMAAA and PHEMAAA / silver nanocomposite film of Example 1 confocal laser scanning microscopy photographs showing that the PHEMAAA / silver nanocomposite films of the present invention exhibit antifouling properties against the three bacteria, as shown here. In Figure 4b a) PS, b) PHEMAAA , c) PHEMAAA / Ag nanocomposite (3 h UV irradiation), and d) PHEMAAA / Ag nanocomposite (represents the heat 3 h, 95 ℃) (5 × 10 7 of CFU / mL 0.0 > 25 C < / RTI > for 24 hours).

Experimental Example 2. Antimicrobial Test

To further investigate the antimicrobial effect, 10 ⁶ CFU / ml of Pseudomonas aeruginosa and Staphylococcus aureus were plated on the PHEMAAA / silver nano composite film for 2 hours, and the number of bacteria attached after a certain period of time was confirmed. The experimental method is as follows.

First, Pseudomonas aeruginosa and Staphylococcus were streaked on LB agar medium and NB (Nutrient Broth) agar medium, respectively, and then cultured in an incubator at 37 캜 for 18 hours with stirring. One bacterial colony was then inoculated into 30 ml of liquid medium. Then, the culture was diluted to 1 x 10 ⁶ CFU / ml using phosphate buffer (0.2M potassium phosphate, 0.07M sodium chloride, 0.07M pH 7.1). The bacterial suspension was dropped onto a polymer sample (25 x 25 mm ² ) (the silver-nanocomposite prepared in Example 1), and then the PET film was stuck on top to allow the bacteria to spread evenly. Take off the film after 3 hours at room temperature, the surfactant (Tween ^® 20, in distilled water (0.2 volume%) and scavengers (quenching agent, 14.6%% sodium thiosulfate by weight, 10.0% by wt thioglycolate) 1: 0.025 by volume 540 ㎕ of the solution containing the bacteria was added to the wells and the bacteria were removed without loss to prevent further antibacterial activity. Finally, the bacteria were plated on agar plates and incubated at 37 캜 for 18 hours. As a result, as shown in Figure 5, in the case of the silver-nanocomposite of the present invention, 99.9% of both bacteria were removed, which is a result of both ultraviolet rays and heat treatment. The PHEMAAA, which was not included, showed no change in the number of living bacteria after 2 hours of contact.

Experimental Example 3. Toxicity Test and Biocompatibility Test

In order to confirm that the PHEMAAA / silver nanocomposite of the present invention can be applied to the biomedical field, the following six tests were performed: i) analysis of the mitochondrial metabolic activity (3- (4,5-dimethylthiazol-2-yl) , 5-diphenyl tetrazolium bromide), ii) neutral red assay, iii) transferase-mediated deoxyuridine triphosphate nick end-labeling, iv) reverse transcription polymerase chain reaction -polymerase chain reaction, v) immunocytochemistry, and vi) cell growth assay.

All of the above tests started with the following procedure. First, human dermal fibroblasts were seeded in TCP (tissue culture polystyrene), PHEMAAA, or PHEMAAA / silver nanocomposite films and then incubated with 10% (v / v) fetal bovine serum and 1% ) Penicillin / streptomycin in Dulbecco's modified Eagle's Medium (DMEM).

(i) MTT assay

First, MTT (3- (4,5-Dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide) analysis was performed to confirm the toxicity of human dermal cells to mitochondrial metabolic activity. MTT solution (2 mg / ml in PBS) was added to human dermal cells and incubated at 37 占 폚 for 4 hours. And replaced with dimethylsulfoxide to dissolve the formazan crystal. The cells were further cultured for 30 minutes while shaking at the same temperature and absorbance was measured at 550 nm. Mitochondrial metabolic activity was determined as the% absorbance of the PS group (TCPS).

(ii) Neutral red dye analysis

Neutral red analysis is performed to assess the viability of human dermal cells and activity is determined based on the degree of absorption of 3-amino-7-dimethylamino-2-methylphenazine hydrochloride. Human dermal cells were washed with PBS and incubated in DMEM containing 50 [mu] g / ml of dye for 3 hours at 37 [deg.] C. After removal of DMEM, a solution of 1% (v / v) acetic acid and 50% (v / v) ethanol was added to extract the dye. After incubation at room temperature for 5 minutes, absorbance was measured at 540 nm.

The results of MTT analysis and neutral red dye analysis are shown in FIG. As shown, the PHEMAAA / silver nanocomposite did not show toxicity to the cells, and the effect on the cell activity was similar to that of TCPS and PHEMAAA, which are commonly used.

(iii) TUNEL analysis

The cytotoxic activity on human dermal cells was confirmed by TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) analysis. Human dermal cells were fixed in a mixed solution of PBS and 4% (v / v) paraformaldehyde and allowed to stand at room temperature for 10 minutes. After adding ethanol and acetic acid (2: 1) mixed solution, the cells were fixed at -20 ° C for 5 minutes and stained using the ApopTag ^® apoptosis detection kit according to the manufacturer's instructions. Cells were stained contrasted with 4,6-diamidino-2-phenylindole and observed with a confocal laser microscope (CLSM, Eclipse 90i, Nikon, Japan, 400X). As a result, as shown in Figs. 7A, 7B, and 7C, in the case of the PHEMAAA / silver nanocomposite film, the effect on cell death was very small (red color) lower than 3% of the whole.

(iv) reverse transcription-polymerase chain reaction (RT-PCR)

Cells grown in each polymer sample of Experimental Example 3 were mixed with TRIzol reagent (Invitrogen, Carlsbad, Calif.). Cellular RNA was then extracted with chloroform and precipitated in isopropanol 80% by volume. The supernatant is removed and the RNA is washed with 75% ethanol and dried. Dissolved in water treated with 0.1% diethyl pyrocarbonate. Next, cDNA was synthesized by performing reverse transcription using 5 μg of pure RNA and SuperScript ^™ IIreversetranscriptase (Invitrogen) according to the manufacturer's instructions and amplified by PCR using the following conditions: denaturation at 94 ° C. for 30 sec; Annealing 58 占 폚, 45 sec, elongation 72 占 폚, 45sec-35 cycles; Final elongation 72 [deg.] C, 45 sec. PCR products were visually confirmed by electrophoresis on 2% (w / v) agarose gels and analyzed using the gel documentation system (Gel Doc 1000, Bio-Rad, Hercules, CA). The results are shown in FIG. As can be seen in Figure 7d, Bax and p53 were not detected in all three samples, indicating that the present nanocomposites are not toxic. In the present invention, genes such as Bax, p53 and PCNA were isolated from RNA and confirmed. P53 is a gene that regulates cell cycle. It acts when DNA is damaged, stops cell cycle and restores damage, or induces cell death in damaged cell. Therefore, when p53 is damaged, it develops into cancer cells. BAX is a protein related to apoptosis. In brief, the expression of Bax increases cell death.

 PCNA (proliferating cell nuclear antigen) is a protein that forms part of a DNA-synthesizing enzyme that acts when DNA replication occurs. PCNA is often used as a marker for proliferating cells.

On the other hand, PCNA associated with cell proliferation activity has been detected, indicating that the complex film enables cell proliferation.

(v) Immunocytochemistry analysis

Human dermal fibroblasts were fixed in 4% (v / v) paraformaldehyde at 25 ° C for 10 min and then washed with PBS. PCNA (proliferating cell nucleus antigen) was reacted with monoclonal antibody (1: 100, Abcam, Cambridge, UK). The cells were then incubated for 1 hour at 25 ° C in PBS containing a rhodamine conjugate secondary antibody (1: 100, Jackson-Immunoresearch, West Grove, PA, USA). Nuclei were stained with 4,6-diamidino-2-phenylindole (DAPI, Vector Laboratories, Burlingame, CA, USA).

(vi) Cell growth assay

For cell growth analysis, proteins used as nutrients of human dermal cells were digested with trypsin, and then counted by a hemacytometer 2 days and 4 days later. As shown in Figure 8, three samples showed almost identical results indicating that silver in the silver-nanocomposite does not inhibit cell growth.

It can be seen from the above Experimental Examples 1 to 3 that the complex of the present invention has antimicrobial activity, antibacterial activity and biocompatibility, and has stability even in water.

While the present invention has been described in connection with what is presently considered to be the preferred embodiments, it is to be understood that the invention is not limited to the disclosed embodiments, but, on the contrary, .

All technical terms used in the present invention are used in the sense that they are generally understood by those of ordinary skill in the relevant field of the present invention unless otherwise defined. The contents of all publications referred to herein are incorporated herein by reference.

Claims (10)

Wherein the polymer comprises poly (2- (2-acetoxyacetoxy) ethyl methacrylate and reduced silver, wherein the reduced silver is incorporated into the polymer.
The polymer-silver nanocomposite according to claim 1, wherein the polymer contains 85 to 99% by weight and the silver contains 0.001 to 5% by weight.
delete Coating a mixture of a biocompatible polymer, poly (2- (2-acetoxyacetoxy) ethyl methacrylate and a silver compound, on a substrate; and
The method of producing a polymer-silver nanocomposite according to claim 1, wherein the coated mixture is treated by electron beam, light irradiation, or heat to reduce the silver compound, and the reduced silver is incorporated in the polymer .
The method for producing a biocompatible polymer / silver nanocomposite according to claim 4, wherein the silver compound is perchloric acid, silver nitrate, or carbonic acid silver.
5. The method for producing a biocompatible polymer / silver nano composite according to claim 4, wherein the light is near ultraviolet light, ultraviolet light or gamma ray.
5. The method of claim 4, wherein the heat treatment is performed at a temperature of 80 to 120 ° C.
An antimicrobial, antifouling, and biocompatible polymer film produced using the biocompatible polymer-silver nanocomposite of claim 1.
An antimicrobial, antifouling, and biocompatible coating material prepared using the biocompatible polymer-silver nanocomposite of claim 1.
A biomedical material having antimicrobial, antifouling and biocompatibility using the biocompatible polymer-silver nanocomposite of claim 1.

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